Inhibitors of HIV integrase are being developed as potential anti-AIDS drugs in collaboration with the Laboratory of Molecular Pharmacology and HIV Drug Resistance Program, CCR, NCI. Lead inhibitor structures have initially been derived from several sources, including three-dimensional pharmacophore searching of the more than 250,000 compounds contained within the NCI's chemical repository. Promising compounds have been systematically explored through chemical synthesis of analogues to determine structure-activity relationships (SAR) responsible for integrase inhibition. Information generated in this fashion has been applied to the design and preparation of new analogues having higher potency, reduced collateral cytotoxicity and greater antiviral protective effects in HIV infected cells. One class of lead structure currently under investigation can be broadly characterized as being of the aryl beta-diketo family. Members of this class have been reported independently to exhibit potent inhibition of HIV integrase in extracellular enzyme assays and to provide good antiviral effects in HIV-infected cells. In collaborative studies with the Rega Institute for Medical Research, Belgium, we have demonstrated that inhibition of HIV replication in whole cells by these compounds occurs in a fashion consistent with inhibition of integrase function. Through the systematic design and synthesis of a large number of aryl beta-diketo analogues, we have developed novel azido-containing aryl beta-diketo variants, which exhibit high integrase inhibitory potency in extracellular assays and provide antiviral effects cells with reduced cytotoxicity in HIV infected. In order to elucidate the manner in which these and other inhibitors interact with integrase-DNA substrate complexes, chemical and photo-activatable affinity labels have been incorporated into high affinity inhibitors. Mass spectral studies are currently ongoing to elucidate sites of covalent attachment of these agents following incubation with the enzyme. In separate studies, collaborative efforts are underway to obtain X-ray structures of inhibitors bound to the HIV integrase enzyme. Information obtained from such X-ray structures should provide a starting point for the computer assisted design of potent new inhibitors.